The aim of this research proposal is to continue an ongoing study to characterize those factors which control the inactivation and reactivation of kidney (NA ions plus K ions) ATPase. The results thus far support the hypothesis that inactivation is mediated in part by a cyclic AMP dependent protein kinase. This kinase is present in kidney microsomes, but absent in purified microsomal (Na ion plus K ion) ATPase. The data suggests that reactivation is mediated by a soluble-protein present in the cellular cytoplasm. The recent observations that vanadate anion is a potent inhibitor will also be studied in order to define whether vanadate inhibition of (Na ions plus K ion) ATPase is independent of the cyclic AMP dependent protein kinase inactivating and cellular soluble-protein reactivating systems, or whether the inhibition of (NA ion plus K ion) ATPase induced by vanadate may also influence the inactivating-reactivating systems. The physiological significance of these interactions will be explored with the use of the isolated perfused kidney. The study will utilize standard biochemical techniques of enzyme purification. The kinetics of the interactions will be studied with cyclic AMP dependent protein kinase alone and with various constituents incorporated with (Na ion plus K ion) ATPase into artificial lipid vesicles. The vanadate experiments will include effects of vanadate binding, vanadate tissue levels and the estimation of kinetics of inhibition. The relationship of "in vitro" effects to "in vivo" effects of cyclic AMP and vanadate inactivation of (Na ion plus K ion) ATPase will be examined with the "in situ" isolated perfused kidney. The perfusion studies will include sodium reabsorption, isobologram analysis of drug interactions and tissue assays of inhibitors and (Na ion plus K ion) ATPase.